One of the grand open challenges in modern science is to image or monitor processes or structures that are very small and operate at a fast time-scale. For example, it is extremely difficult to identify cells or probe molecules and understand the mechanism and dynamics of biological processes at the molecular level with high sensitivity and spatiotemporal resolution, and particularly inside living cells and tissue or liquid. As a result of the wealth of information potentially accessible from such biological targets, there has been a growing demand for imaging tools for biomedical research and medicine. In addition, the continued miniaturization of integrated circuits has led to increasingly small devices. As the distances between components have shrunk, the electric fields generated by these components have increased. These larger electric fields carry the added risk that an electrical short or breakdown effect will occur somewhere in the circuit. If a circuit does not perform to specification, locating troublesome areas can be stymied by the complexity of modern-day circuits; checking each node of the circuit is time-consuming and technically difficult. Accordingly, a need exists to optically measure abnormalities in electric field.
These issues have led to the development of new techniques like magnetic resonance imaging (MRI), ultrasound, positron emission tomography (PET), and optical coherence tomography (OCT). However, these techniques require high costs and some fundamental technological barriers hinder their widespread use.
Optical imaging is a recent technique that utilizes photons as an information source with applications in a wide range of basic science and clinical studies like pharmacology, cellular biology, and diagnostics. For example, semiconductor nanocrystals, small organic dyes or fluorescent proteins are commonly used as optical labels in in vivo optical imaging. (See, e.g., X. Michalet et al., Science 307, 538 (Jan. 28, 2005); B. Dubertret et al., Science 298, 1759 (Nov. 29, 2002); M. K. So, C. Xu, A. M. Loening, S. S. Gambhir, J. Rao, Nat Biotechnol 24, 339 (March, 2006); N. C. Shaner, P. A. Steinbach, R. Y. Tsien, Nat Methods 2, 905 (December, 2005); and B. N. Giepmans, S. R. Adams, M. H. Ellisman, R. Y. Tsien, Science 312, 217 (Apr. 14, 2006), the disclosures of which are incorporated herein by reference.) Moreover, recent advances in fluorescence microscopy alone have profoundly changed how cell and molecular biology is studied in almost every aspect. (For example, see, Lichtman, J. W. & Conchello, J. A. Nat. Methods 2, 910-919 (2005); Michalet, X. et al. Annu. Rev. Biophys. Biomolec. Struct. 32, 161-182 (2003); Jares-Erijman, E. A. & Jovin, T. M. Nat. Biotechnol. 21, 1387-1395 (2003); Bastiaens, P. I. H. & Squire, A., Trends Cell Biol. 9, 48-52 (1999); and Suhling, K., et al., Photochem. Photobiol. Sci. 4, 13-22 (2005), the disclosures of which are incorporated herein by reference.) Indeed, since the cloning of the green fluorescent protein (GFP) from the jellyfish Aequorea victoria a large variety of genetically encoded fluorescent tags have come to be used in in vivo optical imaging. (See, e.g., Prasher, D. C., et al., Gene 111, 229-233 (1992); and Shaner, N. C., et al., Nat Methods 2, 905-909 (2005), the disclosures of each of which are incorporated herein by reference.) They have proven to be particularly important in analyzing a variety of biological processes such as gene expression, and the localization and dynamics of fluorescent-tagged proteins or fluorescent marked cell populations. (See, e.g., Lippincott-Schwartz, J., et al., Nat Rev Mol Cell Biol 2, 444-456 (2001); and Hadjantonakis, A. K., et al., Nat Rev Genet. 4, 613-625 (2003), the disclosures of each of which are incorporated herein by reference.)
However, the ultimate need of characterizing biological targets is largely unmet due to fundamental deficiencies associated with the use of fluorescent agents. For example, fluorescent probes face two major limitations that have a significant impact on the signal strength: 1) dye saturation, because the number of photons emitted by the fluorophore in a given time is restricted by the excited state lifetime, and 2) dye bleaching, which limits the total number of photons per dye. Among these problems, bleaching is perhaps the most significant issue in the application of fluorescent probes, limiting significantly the length of time that biological targets can be studied. In particular, directly tracking the lineage of distinct cell populations in tissue or monitoring the dynamics of molecules within single cells depend critically on long-term photostability. In addition, autofluorescence from tissue organic components due to illumination absorption can severely limit the signal-to-noise ratio. Finally, fluorescence is fundamentally an optically incoherent process, and as a result extracting 3D information from the source is inherently difficult.
Accordingly, a need exists for a new probe for imaging/detecting biological structures and processes that avoids the inherent technological limitations found in the fluorescent imaging techniques of the prior art.